Transient Pax8 expression was reported in mouse islets during gestation, whereas a genome-wide linkage and admixture mapping study highlighted PAX8 as a candidate gene for diabetes mellitus (DM). We sought the significance of PAX8 expression in mouse and human islet biology. PAX8 was induced in gestating mouse islets and in human islets treated with recombinant prolactin. Global gene expression profiling of human and mouse islets overexpressing the corresponding speciesspecific PAX8 revealed the modulation of distinct genetic pathways that converge on cell survival. Accordingly, apoptosis was reduced in PAX8-overexpressing islets. These findings support that PAX8 could be a candidate gene for the study of gestational DM (GDM). PAX8 was genotyped in patients with GDM and gestational thyroid dysfunction (GTD), a pathology commonly found in patients with mutations on PAX8. A novel missense PAX8 mutation (p.T356M, c.1067C>T) was identified in a female diagnosed with GDM and GTD as well as in her father with type 2 DM but was absent in control patients. The p.T356M variant did not alter protein stability or cellular localization, whereas its transactivation activity was hindered. In parallel, a retrospective clinical analysis uncovered that a pregnant female harboring a second PAX8 mutation (p.P25R, c.74C>G) previously reported to cause congenital hypothyroidism also developed GDM. These data indicate that increased expression of PAX8 affects islet viability and that PAX8 could be considered as a candidate gene for the study of GDM. During gestation, maternal metabolic adaptations are essential to ensure the health of the mother and the viability of the fetus. One of the organs that must undergo a profound adaptation during pregnancy is the endocrine pancreas. The islet b-cell mass expands as an adaptive response to the progressive insulin resistance that develops in the pregnant female to favor nutrient accessibility to the fetus (1). This mechanism of b-cell mass expansion includes b-cell hypertrophy, increased b-cell proliferation, and increased prosurvival/antiapoptotic signaling (2). Failure to adapt may result in the development of gestational diabetes mellitus (GDM), with concomitant health issues not only for the mother but also for the fetus. The underlying molecular mechanisms triggering GDM remain largely uncharted (3). Toward identifying these molecular pathways, two independent studies have analyzed the transcriptome landscape of islets isolated from gestating mice (2,4). Of note, one of the most upregulated genes was the transcription factor paired box 8 (PAX8) (2). Historically, PAX8 is known for its essential role in the
BMD3721 to CA). MC was sponsored by the Miguel Servet Program (CPII17_00006) from ISCIII, SA-M was sponsored by the Regional Ministry of Science and Technology of the Board of the Communities of "Castilla-La Mancha" (PREJCCM2016/28) and AT was sponsored by the Regional Government of Madrid/European Regional Development Fund (ERDF) (PEJD-2018-PRE/BMD-9453). M-TG-A was the recipient of a fellowship from the "Ministerio de Educación, Cultura y Deporte" (FPU 13/03308). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
BackgroundThe paired-domain transcription factor paired box gene 6 (PAX6) causes a wide spectrum of ocular developmental anomalies, including congenital aniridia, Peters anomaly and microphthalmia. Here, we aimed to functionally assess the involvement of seven potentially non-canonical splicing variants on missplicing of exon 6, which represents the main hotspot region for loss-of-function PAX6 variants.MethodsBy locus-specific analysis of PAX6 using Sanger and/or targeted next-generation sequencing, we screened a Spanish cohort of 106 patients with PAX6-related diseases. Functional splicing assays were performed by in vitro minigene approaches or directly in RNA from patient-derived lymphocytes cell line, when available.ResultsFive out seven variants, including three synonymous changes, one small exonic deletion and one non-canonical splice variant, showed anomalous splicing patterns yielding partial exon skipping and/or elongation.ConclusionWe describe new spliceogenic mechanisms for PAX6 variants mediated by creating or strengthening five different cryptic donor sites at exon 6. Our work revealed that the activation of cryptic PAX6 splicing sites seems to be a recurrent and underestimated cause of aniridia. Our findings pointed out the importance of functional assessment of apparently silent PAX6 variants to uncover hidden genetic alterations and to improve variant interpretation for genetic counselling in aniridia.
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